Endocytic pathway for surfactant protein A in human macrophages: binding, clathrin-mediated uptake, and trafficking through the endolysosomal pathway

Department of Medicine, Division of Infectious Diseases; Department of Molecular Virology, Immunology, and Medical Genetics; and the Center for Microbial Interface Biology, Ohio State University, Columbus, Ohio Submitted 21 June 2005 ; accepted in final form 13 September 2005 In the noninflamed lung, surfactant protein A (SP-A) acts as an anti-inflammatory molecule through its effects on macrophage (M ) function, modulating cytokine and reactive oxygen and nitrogen intermediate production. The receptors responsible for these effects of SP-A on human M are not clear, although SP-A binding to several proteins has been described. In this study, we demonstrate high-affinity specific binding of SP-A to primary human M . SP-A binding was inhibited by EGTA, indicating calcium dependence. However, mannan did not inhibit SP-A binding, suggesting that binding is mediated by a direct protein-protein interaction that does not involve carbohydrate recognition. Our laboratory has previously shown that SP-A is rapidly endocytosed by human M into discrete vesicles. Although previous work indicates that SP-A is ultimately degraded by murine M over time, the trafficking pathway of SP-A through M after uptake has not been reported and is of potential biological importance. We examined trafficking of SP-A in human M by electron and confocal microscopy and show for the first time that SP-A is endocytosed by primary human M through clathrin-coated pits and colocalizes sequentially over time with the early endosome marker EEA1, late endosome marker lamp-1, and lysosome marker cathepsin D. We conclude that SP-A binds to receptor(s) on human M , is endocytosed by a receptor-mediated, clathrin-dependent process, and trafficks through the endolysosomal pathway. These studies provide further insight into the interactions of SP-A with the M cell surface and intracellular compartments that play important roles in SP-A modulation of lung M biology. surfactant proteins; alveolar macrophage; endocytosis Address for reprint requests and other correspondence: L. S. Schlesinger, Dept. of Internal Medicine, Ohio State Univ., 420 W. 12th Ave., 200 MRF, Columbus, OH 43210 (e-mail: Schlesinger-2{at}medctr.osu.edu )